Synthesis of steroids



SYNTHESIS 6F STEROIDS Josef Fried, New Brunswick, Richard W. Thoma,Somerville, David Perlman, Princeton, and John R. Gerke, FranklinTownship, N. 3., assignors to Olin Mathieson (Ihemical Qorporafion, NewYork, N. Y., a corporation of Virginia No Drawing. Application May 27,1955 Serial No. 511,784

12 Claims. (Cl. 260397.4)

This application is a continuaton-in-part of our copending application,Serial Number 372,798, filed August 6, 1953, now Patent No. 2,753,290,issued July 3, 1956.

This invention relates to the synthesis of valuable steroids.

One object of this invention is the provision of steroids of theprogesterone series having a 7a-hydroxy or 7&- acyloxy group, whichcompounds are. useful for their own physiological action.

The compounds of this invention comprise 7a-hydroxyprogesterone and thecarboxylic acid esters thereof. The preparation of the7oz-hydroxyprogesterone of this invention by the micro-biologicaloxidation of progesterone is disclosed in our Patent No. 2,753,290. Thecarboxylic acid esters of 7a-hydroxyprogesterone of this invention arepreferably prepared from the free hydroxy derivative by reacting thelatter with an acylating agent such as a carbonyl halide or carboxylicacid anhydride as more fully disclosed hereinafter.

Among the compounds of this invention are those of the general formulawherein R is hydrogen or an organic carbonyl radical. Suitable organiccarbonyl radicals include the aliphatic carbonyl radicals, such as thealkanoyl radicals (e. g., lower alkanoyl radicals, such as acetyl,propionyl, butyryl, valeryl, caproyl, and enanthoyl), aroyl radicals (e.g., aromatic hydrocarbon carbonyl radicals, such as benzoyl andnaphthcyl), cycloalkanoyl radicals (e. g., hexahydrobenzoyl), aralkanoylradicals (e. g., a-toloyl and ,B-phenylpropionyl), and heterocycliccarbonyl radicals (e. g., nicotinoyl, furoyl, and 2-thiophenecarbonyl).The preferred organic carbonyl radicals, however, are those ofhydrocarbon carboxylic acids having less than ten carbon atoms, thelower alkanoyls being particularly preferred.

To prepare the steroids of this invention, progesterone is subjected tothe action of the enzymes of a microorganism such as Phycomycesblakesleeanus (Department of Biological Sciences, Purdue University) inan aqueous medium containing a source of nitrogenous factors and anassimilable source of carbon and energy, in the presence of ox gen, andthe 7a-hydroxyprogesterone formed is then recovered. This method is morefully detailed in said Patent No. 2,753,290, and in the examplefollowing.

The 7a-hydroxyprogesterone thus formed can then be acylated by reactingthe 7a-hydroxy steroid with an States Patent acylating agent such as acarbonyl halide or a carboxylic acid anhydride in the presence of abasic agent. Suitable carbonyl halides include the aliphatic carbonylhalides, such as the alkanoyl chlorides (e. g., a lower alkanoylchloride, such as acetyl chloride, propionyl chloride, butyryl chloride,valeryl chloride, caproyl chloride, and enanthoyl chloride), aroylchlorides (e. g., benzoyl chloride and naphthoyl chloride),cycloalkanoyl chlorides (e. g., hexahydrobenzoyl chloride), aralkanoylchlorides (e. g., a-toluyl chloride and p-phenylpropionyl chloride), andheterocyclic carbonyl chlorides (e. g. nicotinoyl chloride, furoylchloride, and 2-thiophenecarbonyl chloride). The preferred carbonylhalides are the hydrocarbon carbonyl chlorides having less than tencarbon atoms, the lower alkanoyl chlorides being particularly preferred.Suitable carboxylic acid anhydrides include the aliphatic carboxylicacid anhydrides, such as the alkanoic anhydrides (e. g., lower alkanoicanhydrides such as acetic anhydride, propionic anhydride, butyricanhydride, valeric anhydride, caproic anhydride, and enanthicanhydride), aromatic carboxylic acid anhydrides (e. g., benzoicanhydride and naphthoic anhydride), cycloalkanoic anhydrides (e. g.,hexahydrobenzoic anhydride), aralkanoic anhydrides (e. g., a-tOlLliCanhydride and B- phenyl-acetic anhydride) and heterocyclic carboXylicacid anhydrides (e. g., nicotinic anhydride, furoic anhydride, and2-thiophenecarboxylic anhydride). The preferred carboxylic acidanhydrides are the hydrocarbon carboxylic acid anhydrides having lessthan nineteen carbon atoms, the lower alkanoic anhydrides beingparticularly preferred. Suitable basic agents are organic bases (e. g.pyridine and collidine) and inorganic bases (e. g the alkali salts oflower fatty acids). is conducted either with an organic base serving asa solvent or in an inert organic solvent such as chloroform, using atleast a stoichiometric amount of acylating agent.

The carboxylic acid esters of 7a-hydroxyprogesterone of this inventionare active materials which possess progestational activity. Thus, thenew steroids of this invention can be administered instead of, and inthe same manner as, progesterone in' the treatment of functional uterinebleeding and amenorrhea. The dosage for such administration is of coursedependent on the relative activity of the particular ester andprogesterone. The 7a-hydroxyprogesterone is also a utilizableintermediate in the preparation of the known steroid, A-dehydroprogesterone, as disclosed in Patent No. 2,753,290, andhereinafter.

The following examples are illustrative of the invention (alltemperatures being in Centigrade):

EXAMPLE 1 7 a-hydroxy progesterone (a) FERMENTATION A medium of thefollowing composition is prepared:

Cornsteep liquor solids 3 NH4H2PO4 3 CaCO 2.5 Soybean oil 22Progesterone 0.50

Distilled water to make one liter.

The acylating reaction q' s 0139- minutesat 120%.] p '(b) ISOLATION 'OFTHE 15BJi DRoXYPRoGESTERONE 4.0-f 0.2 with sulfuric acid, andfiltered bysuction through Seitz filter pads. [The vegetative inoculum used isgrown from stock cultures (lyophili zed vial or agar slant) for 24-72-hours (with or :withou't'successive 24-72hou'r p e riods) in'a medium ofthe following compositionz' 15 g.

co'rnst'eep'liquor solids; 10 g. brown sugar; ,6 g. NaNO MgSQ,,-.7H O;"5g. CaC O 2 g. lard oil; and distilled water to make one liter,the-medium being sterilized by AND THE 7a-HYDR'OXYPROGES TE RONE FORMED'9 litiers of" a culture filtrate obtained as described in (a) byfermentation of 4.85, g.-progesteronejis.extracted;

the contents of the flasks are pooled, adiusted to I with sixZ-litrportions of chloroform. The combined" chloroform extract is filtered,and evaporated to dryness p in vacuo; The residue, weighing about 1.31g., is taken up. in 'm1.'80% aqueous methanol, and the resultingsolution'e xtracted'with five 25 ml. portions of hexane.

The methanol solution' is then evaporateclto dryness, and; j

. the residue (weighing about 1.023 g.) is dissolved with. ,warming in 1ml. of chloroform and 4 ml. o f benzene 'The resultingsolution ischromatographed' on 20 g. of

sulfuric acid-washed alumina. Elution'with 400 ml. of

a mixture of 1 part'of chloroform and 4 parts of henzeneyield's about470 mg. of a 15,8-hYdroxyprogesterone, which after recrystallizationfrom acetone melts at about i M 2.96 .(OH). V 5.90 1. (sat.'CO);I6.06 u(conj. CO); 6.19 ,u (conj. double 7 bond). 7 a

Analysis-Called. for C H O C, 76.33; H, .9.15. Found: C, 76.31; .H,8.90. L g

-' Subsequent elution of the column with equalvolurnes V Vofbenzene-chlorotormyields in the first'l75 ml.-mixed products, andin'the subsequent 800 ml. about mg. p of 7 u-hydroxyprogesterone.. Thelatter, after two crysta1- V lizations .frorn' acetone, melts at about5.8814 (ZO-keto); 6.06 ,1; 6.22 p. (A -3-k6lIOL 7 Ai dhydfoprpgesteronea A"solution of"15 mg. of'the .7arhydroxyprogesterone.5

obtained asdescribedin section (b) of Examplel, in 5 ml. 7

' acetic anhydride of Example 3, the correspondinglower alkanoic esters,such asthepropionate, ,butyrate, and

7 within thescope "4 r a. is acetylated with 1 m1. of pyridine and 1 ml.of acetic anhydride for 18 hours at room temperature. After removal ofthe reagents, in high vacuum the crystalline residue isrecrystallizedfronr acetone. Pure 7a-hydroxyprogesterone acetate has thefollowing properties, M. l.

abQut 38", [a] j +7l' (c, 1.0 in chloroform) its. 237 mp: (e=17,700)

. AnalysisP-QCalcd. for c n o 372.49 C, 74.16;.H,

' v EXAMPLE4,

7a-hydroxyprogesterone enantha'te V Followingtheprocedure of Example?but'substituting 0.01 ml. of n-heptanoic anhydride for the aceticanhydride, 7a-hydroxyprogesterone enanthate is formed.- j EXAMPLES 7 V7a-hydrbgxyprogesterone benzo ate 7 "If 0.02 1. of benioyl chloride. issubstituted for the acetic anhydridefin Example 3,7ot-hydroxyprogesterone 'benzoate is produced In a similar manner, otherprogesterone. canbe prepared. Thus, if. other lower alkanoic anhydrides,such as propionicanhydride, butyric' anhydride, and valeric anhydjr'ide,are substituted for the valerate, are produeed. a

The invention maybe variously of theappended claims. We claim: 7 I 1. Asteroid selected from the ten carbon atoms- 2. 7a-hydroxyprogesterone.

. V 3. A hydrocarbon'carboxylic acid ester. of -7a-hydroxy- 1progesterone, wherein the acid moiety containsless than ten carbonatoms.

4. A lowerfalkanoic acidester of '7a-hydroryprogesterone. V

5. 7a-hydroxyprogesterone acetate. 6. 7m-hydroxyprogesterone enanthate.7. 7a-hydroxyprogesterone benzoate.

8. The process for preparing esters of 7 a-hydroxyproges terone, whichcomprisesreactinggin the presence of a a basicmedium,7a-hydroxyprogesteronewith an acylating of 2.5% KOH in'rnethanol,is refiuxed for one hour-.5.

' 'After cooling, 5 ml. watenis added, and; the methanol is removed invacuo. The aqueous residue'is extracted with chloroform, and thechloroform solution washed with water. After drying overlsodiumfsulfate, the solvent is removed in vacuo, and the residue(weighing about 12.0;

mg.) is dis solved in 0.25' ml..of beniene andjlml. of

hexane for chromatography on alumina. (250mg). Elution of the aluminacolumn with benzene-hexane. (1:4)

yields 'crystalline material; which after two. recrystal:

.25 mg.j of 7ei-hydroicyprogesterone' (M."P. 1227-288) agent selectedfro m the group consisting of .a' carbonyl halide 7 of a hydrocarbon;carb 'oxylic acid containing @less' than ten carboniatomsand a carboxylic' acid anhydride of a hydrocarbon carboxylic' acid containing lessf'than ten carbon atoms, and recovering the ester thus formed. f I r 9.The process, of claim 8 wherein the acylating agent is a' lower alkanoicanhydride.

.10. The process ofclaim8 wherein the acylating agent 1 7' v is aceticanhydride.

infrared comparison withian authentic sample of M-ae- I'hydroprogesterene (M. 145446? shows these two N "products to beidentical. r

11. 'The'processof claim 8 wherein the acylating agent' esters ot7a-hydroxyotherwise embodied group consisting of 7a- 7hydroxyprogesterone and hydrocarbon carboxylic .acid esters thereof;whcreinthe acid, moiety contains less than

1. A STEROID SELECTED FROM THE GROUP CONSISTING OF 7AHYDROXYPROGESTERONEAND HYDROCARBON CARBOXYLIC ACIC ESTERS THEREOF, WHEREIN THE ACID MOIETYCONTAINS LESS THAN TEN CARBON ATOMS.